In the peripheral blood of patients with solid tumors of epithelial origin, studies have identified circulating cells that have characteristics of tumor cells. These cells that are present in the bloodstream of cancer patients (referred to as circulating tumor cells or CTCs) are thought to play an important role in cancer metastasis by breaking loose from a solid tumor, entering the circulation, and then migrating to distant organs to develop secondary tumors. As such, circulating tumor cell (CTC) enumeration and characterization in the blood of cancer patients is useful for cancer prognostic and treatment monitoring purposes. Even though the number of CTCs present in patient blood is very low, robust technologies have been developed to enumerate and characterize CTCs in patient blood samples. CTCs are detectable in the blood of patients with metastatic cancer using a number of different technologies. Since CTCs are rare they need to be enriched from patient blood for accurate enumeration and characterization. Most of the CTC enrichment and identification assays available today are based on enrichment with anti-EpCAM antibodies and subsequent identification using anti-cytokeratin antibodies. An example is the CellSearch® instrument system available from Janssen Diagnostics, Raritan, N.J.
While the presence of CTCs in patients with cancer has been known for over a century, utilization of these rare cells in cancer diagnosis and prognosis was not feasible since methodologies to detect, isolate and characterize CTCs have not been developed until recently. With the development of robust methodologies to enrich, isolate and characterize CTCs in different types of cancers that are found in solid organs, several clinical studies have been conducted to investigate the possible use of CTCs in cancer diagnosis and prognosis. Assays that enumerate CTCs using the Cell Search® system have been developed for use as an aid to monitor patients with metastatic breast, colorectal, and prostate cancers. It has also been shown the potential usefulness of CTC enumeration using the Cell Search™ system for monitoring patients with melanoma, urothelial, and lung cancer.
Factors that limit the utility of CTCs in cancer diagnosis and prognosis are the low abundance and the fragility of the CTCs that may introduce variability in the evaluation of CTCs using different assay platforms. Transportation of blood samples from the site of phlebotomy to another facility is commonly required for CTC enumeration and characterization. During post-phlebotomy blood sample transportation/storage, fragile CTCs may degrade and compromise the accuracy of CTC enumeration and characterization.
There is a growing interest in the use of CTCs in non-invasive diagnosis, prognosis and monitoring of treatment regimens. The low abundance of the CTCs and their fragile nature may introduce variability in the evaluation of CTCs using different assay platforms. This fragile nature of CTCs arises due to the apoptosis of CTCs which begins after separation from the tumor of origin and after removal of blood from a patient. Therefore, it is necessary to address several pre-analytical issues that arise during the time between blood draw and CTC enrichment and characterization in order to effectively preserve the CTCs for analysis. These include delays in blood processing, blood storage temperature, and agitation of the sample during transport and shipment of blood. Such conditions may affect the integrity of already fragile CTCs causing accurate enumeration and characterization of CTCs difficult. As a result, it is important to consider the type of blood collection device and post-phlebotomy conditions while working with CTC samples.
There is thus a need for methods of stabilizing and protecting circulating tumor cells whereby structural integrity is maintained so that shipping and storage is possible with minimal deleterious effect on the circulating tumor cells. There is a further need for such methods where the detrimental effects of aldehyde fixation are avoided.